Chimeric antigen receptor-T cells targeting AFP-GPC3 mediate increased antitumor efficacy in hepatocellular carcinoma

Mingxing Li; Tailin Chen; Rongshi Huang; Yanhui Cen; Feilan Zhao; Rong Fan; Guozhen He

doi:10.1016/j.ajg.2024.12.002

Chimeric antigen receptor-T cells targeting AFP-GPC3 mediate increased antitumor efficacy in hepatocellular carcinoma

Arab J Gastroenterol. 2025 Jan 4:S1687-1979(24)00130-8. doi: 10.1016/j.ajg.2024.12.002. Online ahead of print.

Authors

Mingxing Li¹, Tailin Chen¹, Rongshi Huang², Yanhui Cen³, Feilan Zhao¹, Rong Fan¹, Guozhen He¹

Affiliations

¹ Embryo Formation Teaching and Research Section, Guangxi University of Chinese Medicine, No.13 Wuhe Avenue, Nanning 530200, Guangxi, China.
² Embryo Formation Teaching and Research Section, Guangxi University of Chinese Medicine, No.13 Wuhe Avenue, Nanning 530200, Guangxi, China. Electronic address: [email protected].
³ Embryo Formation Teaching and Research Section, Guangxi University of Chinese Medicine, No.13 Wuhe Avenue, Nanning 530200, Guangxi, China. Electronic address: [email protected].

PMID: 39757079
DOI: 10.1016/j.ajg.2024.12.002

Abstract

Background and study aims: As a novel immunotherapy, chimeric antigen receptor T (CAR-T) cell technology is successful in treating hematologic malignancies, and exhibits potential benefits in partial solid tumors. Therapies targeting one antigen have some weaknesses, and dual-targeted CAR-T cells may be a better option. Alpha-fetoprotein (AFP) and glypican-3 (GPC3) are both highly expressed in hepatocellular carcinoma (HCC) and serve as important markers. Our study aimed to compare the cytotoxicity effect of AFP and GPC3 dual-targeted CAR-T cells on HCC cells in vitro and its therapeutic effects on a SCID xenograft model with those of single-targeted CAR-T cells.

Materials and methods: pLVX lentivirus vectors loaded with AFP CAR, GPC3 CAR, or AFP-GPC3 CAR constructs were transfected into human T lymphocytes. Control T, AFP CAR-T, GPC3 CAR-T, and AFP-GPC3 CAR-T cells were used as effector cells, and HLE (AFP^-GPC3^-), Sh-GPC3-Huh-7 (AFP⁺), Sh-AFP-Huh-7 (GPC3⁺), and Huh-7 (AFP⁺GPC3⁺) cells were used as target cells. After their co-culture for 6 h, the LDH cytotoxicity assay was employed to estimate the cell-killing effects of CAR-T cells on the target HCC cells. SCID mice bearing Huh-7 cell-derived neoplasms were injected with CAR-T cells, after which the pathological changes, CD3ζ expression, and IL-2 and IFN-γ levels in mouse tumor tissues were determined.

Results: AFP and GPC3 were highly expressed in Huh-7 cells. AFP-GPC3 CAR-T cells exerted significant cell-killing effects on the HCC cells that expressed specific targeting antigen molecules (AFP and GPC3). Besides, AFP-GPC3 CAR-T cells better promoted Th cytokine secretion by Huh-7 cells than AFP CAR-T and GPC3 CAR-T cells. In vivo results suggested that AFP-GPC3 CAR-T cells better inhibited the growth of Huh-7 cell (AFP⁺GPC3⁺)-derived neoplasms than AFP CAR-T and GPC3 CAR-T cells.

Conclusion: AFP and GPC3 dual-targeted CAR-T cells showed better anti-tumor effects in HCC than AFP or GPC3 single-targeted CAR-T cells.

Keywords: AFP; Chimeric antigen receptor T cell; GPC3; Hepatocellular carcinoma; Immunotherapy.